Unsupervised correction of gene-independent cell responses to CRISPR-Cas9 targeting

Background: Genome editing by CRISPR-Cas9 technology allows large-scale screening of gene essentiality in cancer. A confounding factor when interpreting CRISPR-Cas9 screens is the high false-positive rate in detecting essential genes within copy number amplified regions of the genome. We have developed the computational tool CRISPRcleanR which is capable of identifying and correcting gene-independent responses to CRISPR-Cas9 targeting. CRISPRcleanR uses an unsupervised approach based on the segmentation of single-guide RNA fold change values across the genome, without making any assumption about the copy number status of the targeted genes. Results: Applying our method to existing and newly generated genome-wide essentiality profiles from 15 cancer cell lines, we demonstrate that CRISPRcleanR reduces false positives when calling essential genes, correcting biases within and outside of amplified regions, while maintaining true positive rates. Established cancer dependencies and essentiality signals of amplified cancer driver genes are detectable post-correction. CRISPRcleanR reports sgRNA fold changes and normalised read counts, is therefore compatible with downstream analysis tools, and works with multiple sgRNA libraries. Conclusions: CRISPRcleanR is a versatile open-source tool for the analysis of CRISPR-Cas9 knockout screens to identify essential genes

Advancement of the German version of the moral distress scale for acute care nurses : a mixed methods study

Aim: Moral distress experienced by nurses in acute care hospitals can adversely impact the affected nurses, their patients and their hospitals; therefore, it is advisable for organizations to establish internal monitoring of moral distress. However, until now, no suitable questionnaire has been available for use in German‐speaking contexts. Hence, the aim of this study was to develop and psychometrically test a German‐language version of the Moral Distress Scale. Design: We chose a sequential explanatory mixed methods design, followed by a second quantitative cross‐sectional survey. Methods: An American moral distress scale was chosen, translated, culturally adapted, tested in a pilot study and subsequently used in 2011 to conduct an initial web‐based quantitative cross‐sectional survey of nurses in all inpatient units at five hospitals in Switzerland's German‐speaking region. Data were analysed descriptively and via a Rasch analysis. In 2012, four focus group interviews were conducted with 26 nurses and then evaluated using knowledge maps. The results were used to improve the questionnaire. In 2015, using the revised German‐language instrument, a second survey and Rasch analysis were conducted. Results: The descriptive results of the first survey's participants (n = 2153; response rate: 44%) indicated that moral distress is a salient phenomenon in Switzerland. The data from the focus group interviews and the Rasch analysis produced information valuable for the questionnaire's further development. Alongside the data from the second survey's participants (n = 1965; response rate: 40%), the Rasch analysis confirmed the elimination of previous deficiencies on its psychometrics. A Rasch‐scaled German version of the Moral Distress Scale is now available for use

Renal artery sympathetic denervation:observations from the UK experience

Background: Renal denervation (RDN) may lower blood pressure (BP); however, it is unclear whether medication changes may be confounding results. Furthermore, limited data exist on pattern of ambulatory blood pressure (ABP) response—particularly in those prescribed aldosterone antagonists at the time of RDN. Methods: We examined all patients treated with RDN for treatment-resistant hypertension in 18 UK centres. Results: Results from 253 patients treated with five technologies are shown. Pre-procedural mean office BP (OBP) was 185/102 mmHg (SD 26/19; n = 253) and mean daytime ABP was 170/98 mmHg (SD 22/16; n = 186). Median number of antihypertensive drugs was 5.0: 96 % ACEi/ARB; 86 % thiazide/loop diuretic and 55 % aldosterone antagonist. OBP, available in 90 % at 11 months follow-up, was 163/93 mmHg (reduction of 22/9 mmHg). ABP, available in 70 % at 8.5 months follow-up, was 158/91 mmHg (fall of 12/7 mmHg). Mean drug changes post RDN were: 0.36 drugs added, 0.91 withdrawn. Dose changes appeared neutral. Quartile analysis by starting ABP showed mean reductions in systolic ABP after RDN of: 0.4; 6.5; 14.5 and 22.1 mmHg, respectively (p < 0.001 for trend). Use of aldosterone antagonist did not predict response (p < 0.2). Conclusion: In 253 patients treated with RDN, office BP fell by 22/9 mmHg. Ambulatory BP fell by 12/7 mmHg, though little response was seen in the lowermost quartile of starting blood pressure. Fall in BP was not explained by medication changes and aldosterone antagonist use did not affect response

Abstracts from the NIHR INVOLVE Conference 2017

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Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

Effect of remote ischaemic conditioning on clinical outcomes in patients with acute myocardial infarction (CONDI-2/ERIC-PPCI): a single-blind randomised controlled trial.

BACKGROUND: Remote ischaemic conditioning with transient ischaemia and reperfusion applied to the arm has been shown to reduce myocardial infarct size in patients with ST-elevation myocardial infarction (STEMI) undergoing primary percutaneous coronary intervention (PPCI). We investigated whether remote ischaemic conditioning could reduce the incidence of cardiac death and hospitalisation for heart failure at 12 months. METHODS: We did an international investigator-initiated, prospective, single-blind, randomised controlled trial (CONDI-2/ERIC-PPCI) at 33 centres across the UK, Denmark, Spain, and Serbia. Patients (age >18 years) with suspected STEMI and who were eligible for PPCI were randomly allocated (1:1, stratified by centre with a permuted block method) to receive standard treatment (including a sham simulated remote ischaemic conditioning intervention at UK sites only) or remote ischaemic conditioning treatment (intermittent ischaemia and reperfusion applied to the arm through four cycles of 5-min inflation and 5-min deflation of an automated cuff device) before PPCI. Investigators responsible for data collection and outcome assessment were masked to treatment allocation. The primary combined endpoint was cardiac death or hospitalisation for heart failure at 12 months in the intention-to-treat population. This trial is registered with ClinicalTrials.gov (NCT02342522) and is completed. FINDINGS: Between Nov 6, 2013, and March 31, 2018, 5401 patients were randomly allocated to either the control group (n=2701) or the remote ischaemic conditioning group (n=2700). After exclusion of patients upon hospital arrival or loss to follow-up, 2569 patients in the control group and 2546 in the intervention group were included in the intention-to-treat analysis. At 12 months post-PPCI, the Kaplan-Meier-estimated frequencies of cardiac death or hospitalisation for heart failure (the primary endpoint) were 220 (8·6%) patients in the control group and 239 (9·4%) in the remote ischaemic conditioning group (hazard ratio 1·10 [95% CI 0·91-1·32], p=0·32 for intervention versus control). No important unexpected adverse events or side effects of remote ischaemic conditioning were observed. INTERPRETATION: Remote ischaemic conditioning does not improve clinical outcomes (cardiac death or hospitalisation for heart failure) at 12 months in patients with STEMI undergoing PPCI. FUNDING: British Heart Foundation, University College London Hospitals/University College London Biomedical Research Centre, Danish Innovation Foundation, Novo Nordisk Foundation, TrygFonden

Improving gene activated matrices for orthopaedic bone regeneration: alternative biomaterials and predicted mRNA structure based BMP2 signal peptide optimisation

Critical and non-union defects are bone injuries that will not fully heal spontaneously, with existing treatments frequently unable to improve outcomes. These defects represent an increasing burden on health systems world-wide and there is an unmet clinical need for improved treatments. Gene-activated matrices (GAMs) combine a biomaterial and a gene therapy, attempting to improve performance by providing multiple osteogenic stimuli simultaneously. This project aims to develop a GAM from a self-assembling peptide (SAP) P11-4 with known hard tissue regeneration capabilities and plasmid vectors encoding the osteogenic growth factor bone morphogenetic protein 2 (BMP2), resulting in an injectable GAM that promotes a multifaceted approach to osteogenesis. New reporter vectors were created and transfection optimisation experiments performed to facilitate the P11-4 GAM work. Transfection from P11-4 GAMs proved difficult and several experiments investigating vector polyplex size and hydrogel porosity were performed to elucidate the mechanisms. A cross-linked gelatine cryogel was then developed as a passive scaffold to allow continued assessment of the plasmid vectors, demonstrating suitable porosity, sterility and biocompatibility. An in silico signal peptide (SP) screening approach based on predicted mRNA structure was designed to identify new SPs to improve BMP2 secretion. SPs from the TGF-β superfamily were screened, with the top five candidates and two controls taken forward to in vitro testing in two cell lines. The approach showed weak predictive power in one line, but none of the tested sequences were able to induce a significant increase in BMP2 secretion or induce osteogenesis. These results suggest that substantial further work would be required to create a P11-4-based GAM, the results presented here indicate that porosity and bioactivity are particularly important. The in silico SP optimisation approach successfully identified several SPs never before used for SP optimisation, but required improvement to generate biologically significant predictions

Guidance for captioning rich media: Institutional approaches to supporting disabled learners

This guidance does not represent legal advice but a summary of likely best practices achievable with current technologies, resources and workflows. This guidance is endorsed by the Digital Accessibility Working Group | December 2020.We recognise that digital accessibility crosses different areas of expertise so all theterminology will not be familiar to everyone. Please note we have provided a glossary in the appendix.Also note that while the guidance refers to lecture capture as an example throughout, the actions and principles can be applied across any rich media.<br/

Impairments in impulse control in animal models transgenic for the human FTPD-17 tauV337M mutation are exacerbated by age

Abnormalities in microtubule-associated tau protein are a key neuropathological feature of both Alzheimer's disease and many frontotemporal dementias (FTDs), including hereditary FTD with Parkinsonism linked to chromosome 17 (FTDP-17). In these disorders, tau becomes aberrantly phosphorylated, leading to the development of filamentous neurofibrillary tangles in the brain. Here we report, in a longitudinal ageing study, the sensorimotor and cognitive assessment of transgenic mice expressing the human tauV337M (‘Seattle Family A’) FTDP-17 mutation, which we have previously shown to demonstrate abnormalities in brain tau phosphorylation. The data indicated highly specific effects of transgene expression on the ability to withhold responding in a murine version of the 5-choice serial reaction time task, behaviour consistent with deficits in impulse control. Ageing exacerbated these effects. In young tauV337M mice, increased impulsivity was present under task conditions making inhibition of premature responding more difficult (longer inter-trial intervals) but not under baseline conditions. However, when older, the tauV337M mice showed further increases in premature responding, including under baseline conditions. These impulse control deficits were fully dissociable from sensorimotor or motivation effects on performance. The findings recapitulate core abnormalities in impulsive responding observed in both frontal variant FTD and FTDP-17 linked to the tauV337M mutation in humans